In the intestinal epithelium, the balance between proliferation and apoptosis within the crypts of Lieberkuhn maintains villus height and crypt depth; disruption of this balance leads to cancer. Within the crypts, stem cells produce both absorptive and secretory cells. Several genes have been implicated in this process, but delineation of an ordered hierarchy for control of intestinal epithelial differentiation is lacking. One gene in this hierarchy, Math1, is required for secretory lineage development; mice lacking Math1, produce absorptive enterocytes but none of the secretory lineages. This proposal will test the hypotheses that Math1- dependent secretory lineages are required locally for proliferative homeostasis in the intestine, and that delineation of the genetic pathway for secretory lineage differentiation will clarify the mechanism of intestinal homeostasis. First I will characterize the genetic pathway for intestinal differentiation by microarray-based expression profiling of Math1 -/- crypts and crypts null for Gfi1, a putative Math1 target gene also shown to have a role in intestinal epithelial development. Experiments in transfected cells will define direct gene targets of these transcription factors. Next, I will develop an in vivo lentiviral-siRNA system to ascertain the phenotype of Notch1 loss in the intestine, and ascertain its position in the hierarchy of intestinal differentiation. In Aim 3, I will assess the effect of azoxymethane to produce tumors and aberrant crypt foci in mice in which the ileum and colon contains Math1 -/- crypts that produce no secretory cells. The data generated from these experiments will form the basis of an independent research program to investigate intestinal development and function. In addition, the aims represent distinct approaches to understanding intestinal biology, and they will inform one another while not being interdependent for success. I will be mentored by Dr. Susan Henning, a leader in the field of intestinal biology and development, with a scientific advisory committee composed of experts in relevant fields. A rich intellectual environment and extensive resources are available to the PI.